The present invention relates to methods and apparatus for maintaining the activity of an enzyme electrode, and more particularly to methods and apparatus for maintaining an enzyme electrode in activated condition while it is provided in a concentration measuring apparatus for measuring the concentration of a test substance in a test solution based upon an oxidation reaction or a reduction reaction of the test substance in a surfacial region of the enzyme electrode.
It is known that a physiologically active substance is capable of selectively detecting a very complicated organic compound, a protein or the like with high sensitivity. With attention directed to this characteristic, research has been made in various biosensors.
A typical biosensor (hereinafter referred to as an enzyme electrode) having an electrode unit and a physiologically active substance fixed thereon is proposed. The enzyme electrode is used for detecting the existence of a test substance, the relative or active quantity of the test substance and the like based on an electrical signal output from the electrode unit corresponding to the biological reaction of the test substance, under the condition that a predetermined forward bias is applied to the electrode unit. For example, the electrode unit generally has a working electrode made of platinum having high activity and a counter electrode made of silver having a high stability. When glucose as the test substance in the test solution is to be measured, the enzyme electrode generally has an arrangement wherein a hydrogen peroxide penetration membrane, an enzyme-immobilized membrane including glucose oxidase as a physiologically active substance, and a diffusions-limiting membrane are laminated in this order. When the enzyme electrode is employed, glucose is oxidized in the presence of glucose oxidase so as to generate gluconic acid and hydrogen peroxide. Then the generated hydrogen peroxide reaches the surface of the electrode unit through a hydrogen peroxide penetration membrane. The electrode unit outputs an electrical signal corresponding to the quantity of hydrogen peroxide that reaches it. The relative or active quantity of glucose in test solution is detected based on the electrical signal. In this case, when the activity of the platinum electrode is lowered, the level of the electrical signal is remarkably lowered. When the platinum electrode is kept as it is for a long time period, an oxidized layer which interferes with electrical signals is formed on the platinum electrode so as to lower the activity of the platinum electrode.
The same disadvantages as above arise for an enzyme electrode which is used for measuring a concentration of a test substance other than glucose.
It is proposed that a reviving voltage, which has a polarity which is reverse with respect to the polarity of a voltage for concentration measurement, is applied to the electrode unit so as to remove the oxidized layer on the electrode unit thereby to enable a concentration measurement with high sensitivity.
A disadvantage arises in that the oxidized layer cannot be perfectly removed even when the reviving voltage is applied to the electrode unit due to an increase in the thickness of the oxidized layer on the electrode unit. The increase in thickness is determined based on lengthening of the time interval between concentration measurements. In this case, when concentration measurements are carried out with a relatively short time interval, the thickness of the oxidized layer which remains after the revising voltage is applied to the electrode unit is gradually decreased at every concentration measurement so as to improve the activity of the electrode unit. As a result, drift appears in the output electrical signals so as to lower the accuracy in the concentration measurement (see FIG. 9).
To reduce the disadvantage above-mentioned, it is necessary that the reviving voltage is raised or the time period for applying the reviving voltage is lengthened. This causes an increase in the quantity of the generated substance, i.e. hydrogen, hydrogen ions and the like, which is generated on the electrode unit by applying the reviving voltage so as to increase the diffusion current which is caused by the generated substance generated. As a result, the necessary time period for the decreasing of the diffusion current to a predetermined threshold value is reduced. The disadvantage arises that the time period is lengthened between performing the reviving operation for the electrode unit to when it is possible to start a measuring operation for a test substance.
The same disadvantages as above arise for an enzyme electrode having a reference electrode in addition to the working electrode and the counter electrode.